A immunosensor for the diagnosis of canine distemper virus infection using SPR and EIS Caroline Rodrigues Basso, a Claudia de Camargo Tozato, b Marcela Cristina Mendes Ribeiro, b Jo~ ao Pessoa Araujo Junior b and Valber A. Pedrosa * a Canine distemper virus (CDV) is a viral disease that affects dogs and many other carnivores. Clinical diagnosis of CDV is difficult due to the broad spectrum of signs that may be confounded with other respiratory and enteric diseases of dogs. Laboratory analysis is required to diagnose suspected cases. In this study, surface plasmon resonance (SPR) and electrochemical impedance spectroscopy (EIS) methodologies were developed for the detection of canine distemper virus simultaneously. The assay exhibited high specificity, as all the negative controls were not mistakenly detected. The CDV concentration was determined from successive injections into the apparatus, with a linear range from 1.1 to 116.0 ng mL 1 . The system exhibited good reproducibility with 4.5% variation between runs after regeneration of the coated surface with a solution of 0.1 M glycine–HCL (pH 3.0). The capacitance and resistance values of the modified interface were calculated from EIS data using an equivalent circuit. It was possible to measure CDV in highly concentrated viruses with good specificity and reproducibility. 1 Introduction Canine distemper is a highly contagious and potentially fatal disease of dogs and many other carnivores. Canine distemper is caused by the canine distemper virus, a Morbillivirus belonging to the Paramyxoviridae family, and it is a highly contagious disease with a worldwide distribution. 1 The virus can be trans- mitted by aerosols or contact with oral, respiratory, and ocular uids, or other virus-containing exudates. Eradication of CDV is considered impossible because of the virus's wide host range, which includes terrestrial carnivores and marine mammals. Use of a disease-specic laboratory diagnosis is essential because clinical signs are nonspecic. Thus, when dealing with this highly contagious illness, specic diagnosis can prevent the spread of the disease between contacts, mainly in kennels and shelters, as well as facilitate the segregation of affected animals in hospitals, thus avoiding contamination of other internees. The high mortality rates of this viral disease necessitate the acceleration of diagnostic procedures to quarantine infected individuals and start the appropriate treatment early. 2 Although there is no specic therapy for the deadly disease, quick and reliable detection methods are extremely critical and essential for measuring the virus at very low levels, not only to save lives but also to save analysis time and money. 3 There are several techniques that can be used to test for CDV infection in dogs, including immunouorescence, 4 immunohistochemistry, 5 serology, 6 indirect uoroimmunoassay, 7 histopathology, 8 and reverse transcription-polymerase chain reaction (RT-PCR). 9 However, each of these tests has limitations. Immunouores- cence can only be performed on fresh-frozen tissue and the test is most reliable within the rst 3 weeks of infection in acute disease. Immunohistochemistry is a highly sensitive and specic method for the detection of the CDV antigen in tissue obtained postmortem, but it is suitable only within limits for the diagnosis of distemper in living animals and is dependent on specic antibody-xation parameters. Histopathology is not specic and normally requires a conrmatory test. Cell culture is time-consuming because it can provide results only in a few days, but specimen quality and origin can limit the effectiveness of this technique. RT-PCR is a technique with high analytical sensitivity, but this varies with the type of sample, RNA extrac- tion method and choice of primers. However, when compared with other techniques for detecting viruses, its advantages are many like, for example, the achievement of rapid and early diagnosis. However, these methods require multi-step pro- cessing of samples, live virus or animal/cell cultures, as well as considerable labor, time and expense, which makes them unsuitable. Therefore, it remains a challenge to nd new approaches that improve the rapidity, simplicity, selectivity, and sensitivity of detection of infectious viral diseases. Recently, biosensors have been widely applied in various elds such as pathogenic analysis, food safety, environmental monitoring, and clinical diagnostics. 10–14 Efforts have been made by different groups to improve the selectivity and sensi- tivity of the biosensors and explore new concepts in a Department of Chemistry and Biochemistry, Institute of Bioscience, UNESP-Botucatu, SP 18618-000, Brazil. E-mail: vpedrosa@ibb.unesp.br; Tel: +55(14) 3880-0576 b Department of Microbiology and Immunology, Institute of Bioscience, UNESP- Botucatu, SP 18618-000, Brazil Cite this: Anal. Methods, 2013, 5, 5089 Received 2nd July 2013 Accepted 15th July 2013 DOI: 10.1039/c3ay41087k www.rsc.org/methods This journal is ª The Royal Society of Chemistry 2013 Anal. 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